摘要
目的探究隐丹参酮(CPT)对脂多糖(LPS)诱导的小胶质细胞功能的作用机制。方法体外培养小胶质HMC3细胞,分为对照组、LPS组、不同浓度CPT组,细胞计数试剂盒-8(CCK-8)与酶联免疫吸附试验(ELISA)分别检测细胞活力和炎症因子表达水平,筛选出合适的CPT作用浓度。随后将细胞分为Con组、LPS组、CPT组、AG490组、抑制剂组和激活剂组,5-乙炔基-2’脱氧尿嘧啶核苷(EdU)、Hoechst 33258染色法分别检测各组细胞增殖率、凋亡率;蛋白免疫印迹(WB)法检测细胞周期蛋白D1(CyclinD1)、半胱氨酸蛋白酶-3(Caspase-3)及JAK2/STAT3通路相关蛋白表达水平。结果5μmol·L^(-1)CPT为最适浓度。LPS组细胞增殖率和CyclinD1蛋白表达水平低于Con组(P<0.05),凋亡率、Caspase-3、p-JAK2和p-STAT3蛋白表达水平高于Con组(P<0.05);与LPS组比较,CPT组和AG490组扭转了上述LPS作用(P<0.05);与CPT组比较,AG490增强、C-A1削弱了CPT对HMC3细胞的作用(P<0.05)。结论CPT负调控JAK2/STAT3通路促进LPS诱导的小胶质HMC3细胞的增殖,抑制其凋亡。
Objective To investigate the mechanism of cryptotanshinone(CPT)on microglia function induced by lipopolysaccharide(LPS).Methods Microglia HMC3 cells were cultured in vitro and divided into the control group,LPS group and different concentrations of CPT group.Cell counting kit-8(CCK-8)and enzyme-linked immunosorbent assay(ELISA)were used to detect cell viability and inflammatory factor expression levels to screen out the appropriate action concentration of CPT.Then the cells were divided into the control group,LPS group,CPT group,AG490[Janus kinase 2/signal transducer and activator of transcription 3(JAK2/STAT3)signaling pathway inhibitor.]group,inhibitor group and activator(JAK2/STAT3 signaling pathway agonist C-A1)group.5-Ethynyl-2'-deoxyuridine(EdU)and Hoechst 33258 staining were used to detect the cell proliferation rate and apoptosis rate in each group.Western blot(WB)was used to detect the expression levels of CyclinD1,Caspase-3 and JAK2/STAT3 pathway-related proteins(p-JAK2 and p-STAT3 protein).Results 5μmol·L^(-1)CPT was the optimal concentration.The cell proliferation rate and CyclinD1 protein expression level in the LPS group were lower than those in the control group(P<0.05),and the apoptosis rate and Caspase-3,p-JAK2 and p-STAT3 protein expression levels were higher than those in the control group(P<0.05).Compared with the LPS group,the above LPS effects were reversed in CPT group and AG490 group(P<0.05).Compared with CPT group,AG490 enhanced the effect of CPT on HMC3 cells,while C-A1 attenuated the effect of CPT(P<0.05).Conclusion CPT negatively regulates the JAK2/STAT3 pathway to promote the proliferation and inhibit the apoptosis of microglial HMC3 cells induced by LPS.
作者
杨鹏
王梦梦
宋梅梅
张晓宇
王淑燕
YANG Peng;WANG Mengmeng;SONG Meimei;ZHANG Xiaoyu;WANG Shuyan(Department of Laboratory Medicine,the Second Central Hospital of Baoding,Baoding 072750,China;Department of Pharmacy,the Second Central Hospital of Baoding,Baoding 072750,China;Department of Internal Medicine,Staff Hospital of Lekai Group Co.,Ltd.,Baoding 071000,China)
出处
《长春中医药大学学报》
2023年第8期875-880,共6页
Journal of Changchun University of Chinese Medicine
基金
保定市2021年度科技计划项目(2141ZF016)
河北省中医药管理局科研计划项目(2021276)。
关键词
阿尔茨海默病
小胶质细胞
隐丹参酮
脂多糖
增殖
凋亡
Alzheimer's disease
microglia
cryptotanshinone
lipopolysaccharide
proliferation
apoptosis
